The Next-Generation Oral Selective Estrogen Receptor Degrader Camizestrant (AZD9833) Suppresses ER+ Breast Cancer Growth and Overcomes Endocrine and CDK4/6 Inhibitor Resistance

Abstract

Oral selective estrogen receptor degraders (SERD) could become the backbone of endocrine therapy (ET) for estrogen receptor-positive (ER+) breast cancer, as they achieve greater inhibition of ER-driven cancers than current ETs and overcome key resistance mechanisms. In this study, we evaluated the preclinical pharmacology and efficacy of the next-generation oral SERD camizestrant (AZD9833) and assessed ER-co-targeting strategies by combining camizestrant with CDK4/6 inhibitors (CDK4/6i) and PI3K/AKT/mTOR-targeted therapy in models of progression on CDK4/6i and/or ET. Camizestrant demonstrated robust and selective ER degradation, modulated ER-regulated gene expression, and induced complete ER antagonism and significant antiproliferation activity in ESR1 wild-type (ESR1wt) and mutant (ESR1m) breast cancer cell lines and patient-derived xenograft (PDX) models. Camizestrant also delivered strong antitumor activity in fulvestrant-resistant ESR1wt and ESR1m PDX models. Evaluation of camizestrant in combination with CDK4/6i (palbociclib or abemaciclib) in CDK4/6-naive and -resistant models, as well as in combination with PI3Kαi (alpelisib), mTORi (everolimus), or AKTi (capivasertib), indicated that camizestrant was active with CDK4/6i or PI3K/AKT/mTORi and that antitumor activity was further increased by the triple combination. The response was observed independently of PI3K pathway mutation status. Overall, camizestrant shows strong and broad antitumor activity in ER+ breast cancer as a monotherapy and when combined with CDK4/6i and PI3K/AKT/mTORi.

Significance: Camizestrant, a next-generation oral SERD, shows promise in preclinical models of ER+ breast cancer alone and in combination with CDK4/6 and PI3K/AKT/mTOR inhibitors to address endocrine resistance, a current barrier to treatment.

Figure 1.

Camizestrant (AZD9833) is a selective ER degrader and pure antagonist. A, Chemical structure of camizestrant. B, The indicated cell lines were treated with 100 nmol/L of the indicated compound for 48 hours. Levels of ERα were assessed by Western blotting and normalized to an untreated control and fulvestrant. Each point represents an independent experiment. C, MCF7 or CAMA-1 cells were treated with DMSO, 1 nmol/L estradiol, or 100 nmol/L of the indicated compound + 1 nmol/L estradiol for 24 hours. RNA expression was assessed by RNA sequencing. Data represent z-scores of normalized gene expression for genes in an ER activity signature. D, MCF7 and CAMA-1 cells were treated with the indicated concentration of the indicated compound for 7 days. Cell number was estimated with a Sytox Green assay normalized to an untreated control on the day of treatment (0.0) and an untreated control on day 7 after treatment (1.0). Data points represent the mean from three independent experiments performed in triplicate ± SD. E, Ishikawa cells were treated with the indicated concentration of fulvestrant or camizestrant, or 100 nmol/L AZD9496 for 24 hours, and ERα and PgR expressions were determined by Western blot. DMSO, dimethyl sulfoxide; PR, progesterone receptor.

Figure 2.

Binding and activity of camizestrant in clinically relevant ERα mutations. A, The pIC₅₀ value of fulvestrant and camizestrant to displace a fluorescent ER ligand from wild-type, D538G, Y537N, E380Q, Y537C, S463P, or Y537S mutant purified ERα ligand-binding domain. Points represent independent experiments. B, MCF7 cells expressing WT or Y537S ERα were grown for 7 days in 5% FBS. Growth inhibition was estimated with a Sytox Green assay normalized to an untreated control on day 0 (0%) and an untreated control on day 7 of treatment (100%). Data points represent the mean from two independent experiments carried out in duplicate. Fulvestrant and camizestrant inhibited the proliferation of both WT and Y537S ERα-expressing MCF7 cells in a concentration-dependent manner. The table shows pIC₅₀ values from independent experiments. C, MCF7 cells expressing WT or Y537S ERα were treated with the indicated concentration of fulvestrant or camizestrant for 72 hours, and ERα was determined by Western blot. Fulvestrant and camizestrant showed concentration-dependent inhibition of PgR expression (normalized to an untreated control) in MCF7 cells expressing both WT and Y537S ERα. D, Camizestrant dose–response in the long-term estrogen-deprived ESR1wt PDX model, HBXF079-LTED. Statistical analysis was performed by one-tailed, unequal variance t test versus log (change in tumor volume) compared with vehicle control at the final day of treatment. E, ER degradation measured by Western blot from tumors taken at the end of the efficacy dosing period. F, In the ESR1m D538G PDX CTC-174 model, camizestrant demonstrated antitumor activity in a dose-dependent manner, with maximal antitumor activity at 10 mg/kg. Efficacy correlated with ER degradation measured by Western blot from tumors taken at the end of the efficacy dosing period. Statistical analyses were performed by one-tailed, unequal variance t test versus log (change in tumor volume) compared with vehicle control at the final day of treatment. G, ER degradation measured by Western blot from tumors taken at the end of the efficacy dosing period. NS, not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001. pIC₅₀, negative log of the IC₅₀ (half-maximal inhibitory concentration) value when converted to mol/L.

Figure 3.

Camizestrant has superior in vivo activity to fulvestrant in ESR1wt and ESR1m PDX models (1). A, Waterfall plot representing the growth of 28 of ER⁺ breast cancer PDX treated with camizestrant 10 mg/kg daily; bars are colored according to ESR1 mutational status and other genomic characteristics annotated at the top. The percentage change calculated from the initial volume at day of treatment is shown. Data represent mean ± SE of the mean. B, Waterfall plot representing the growth of 25 PDX treated with fulvestrant 5 mg/kg weekly. N/A denotes models where the head-to-head arm was not available. The percentage of change calculated from the initial volume at day of treatment is shown. Data represent mean ± SE of the mean. C, Pie chart indicates the proportions of PDX models sensitive or insensitive to camizestrant and/or fulvestrant. The antitumor response of camizestrant versus fulvestrant monotherapy is represented as the percentage of tumor change compared with the initial tumor volume, benchmarked to vehicle changes. D, Correlation of fulvestrant PDX antitumor response (y-axis) versus camizestrant (x-axis) in 25 PDX, represented as the percentage of tumor volume change compared with the initial tumor volume. Data represent mean. The boxes indicate the percentage change from baseline ≤100%; the percentage of models sensitive only to fulvestrant or camizestrant, or sensitive to both is represented in each box.

Figure 4.

Camizestrant has superior in vivo activity to fulvestrant in ESR1wt and ESR1m PDX models (2). A, Characteristics of ER⁺ breast cancer models used. B, Change in ER pathway gene activation after treatment, expressed as change in ER pathway gene score and in cell-cycle G₁–S checkpoint genes. See Supplementary Methods for details. Statistical analysis comparing fulvestrant and camizestrant was done using one-way analysis of covariance (n ≥ 4 animals per group). Models shown in bold (x-axis) are fulvestrant sensitive; those in regular type are fulvestrant resistant. ***, P < 0.001; ****, P < 0.0001. Amp, amplification; CCND1, cyclin D1; Del, deletion; MET, metastasis; Mut, mutation; PIK3CA, phosphatidylinositol 3-kinase subunit α; PR, progesterone receptor; PRIM, primary; RESIST, resistant; RB1, retinoblastoma gene; SENS, sensitive. Patient treatment reported: Be, bevacizumab; Ch, chemotherapy; E, exemestane; Ev, everolimus; F, fulvestrant; I, investigational; L, letrozole, T, tamoxifen; X, radiotherapy.

Figure 5.

Enhanced efficacy of camizestrant in combination with PI3K/AKT/mTOR inhibitors as doublets in CDK4/6-sensitive and -resistant models (1). A–D, Combination of camizestrant with PI3Kα inhibitor alpelisib (A), mTOR inhibitor everolimus (B), AKT inhibitor capivasertib (C), or CDK4/6 inhibitor palbociclib (D) delivers enhanced efficacy compared with monotherapy in D538G ESR1m PDX CTC-174. Statistical analysis was performed by one-tailed, unequal variance t test versus log (change in tumor volume) compared with vehicle control at the final day of treatment. E, Relative tumor volume plots of ST3632 PDX model treated with oral camizestrant at 10 mg/kg daily, oral palbociclib at 50 mg/kg daily, and oral abemaciclib 50 mg/kg daily, and with camizestrant + abemaciclib and camizestrant + palbociclib. Statistical analysis was performed by one-tailed, unequal variance t test versus log (change in tumor volume) at the final day of treatment. F,In vivo combination of camizestrant at 10 mg/kg daily with palbociclib 50 mg/kg, abemaciclib 50 mg/kg, and capivasertib 130 mg/kg in PDX ST1799 dosed for 40 days (gray area). For clarity, the graph is divided into four subgraphs due to the large number of treatment arms; where they appear, the vehicle, camizestrant, and palbociclib arms are the same in each subgraph. CDK, cyclin-dependent kinase. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant.

Figure 6.

Enhanced efficacy of camizestrant in combination with PI3K/AKT/mTOR inhibitors as doublets in CDK4/6i-resistant models (2). A, 28-, 35- or 42-day efficacy studies using several ER⁺ breast cancer PDX harboring/not harboring alterations in PIK3CA/AKT/PTEN. Dark blue, mutations; light blue, deletions; orange, fusions. The rate of growth for each animal is estimated on the basis of fitting each tumor's growth curve to an exponential model: log₁₀(tumor volume) = a + b·time + error, where a and b correspond to the log initial volume and growth rate, respectively. The model assumes that the error terms are normally distributed. Tumor volumes less than 15 mm³ were replaced with a minimum value of 15 mm³. This growth rate summary metric was then used for statistical analysis to compare treatments with a user-specified reference group. Tumor growth inhibition was used to plot a heat map. Designed dosing: oral palbociclib 50 mg/kg daily, subcutaneous fulvestrant 5 mg weekly, oral camizestrant 10 mg/kg daily, oral capivasertib 130 mg/kg BID 4 days on/3 days off. Statistical analysis was performed by one-tailed, unequal variance t test versus log (change in tumor volume) at the final day of treatment. B, 28-, 35- or 42-day efficacy studies used in A; relative tumor volume plots displaying arms: control, standard-of-care hormone therapy + CDK4/6 inhibitor (fulvestrant + palbociclib), or triplet combination of hormone therapy + CDK4/6 inhibitor + AKTi (camizestrant + palbociclib + capivasertib). Designed dosing: oral palbociclib 50 mg/kg daily, subcutaneous fulvestrant 5 mg weekly, oral camizestrant 10 mg/kg daily, oral capivasertib 130 mg/kg BID 4 days on/3 days off. Statistical analysis was performed by one-tailed, unequal variance t test versus log (change in tumor volume) at the final day of treatment. C, Camizestrant fits centrally in the overall landscape of breast cancer as a backbone endocrine therapy. Estrogens (e.g., E2) bind to ERα, leading to its dimerization and translocation to the nucleus, where ERα dimers bind to coactivators to form transcriptionally active ERα complexes. Activated complexes regulate gene transcription in the nucleus or activate kinases in the cytoplasm to drive cell proliferation. Mutations in the ligand-binding domain of ESR1 drive resistance in advanced ER⁺ breast cancer and act independently of estrogens to activate transcription. Camizestrant is a next-generation SERD for the treatment of ER⁺ breast cancer, acting as a pure ER antagonist and selective ERα degrader. Camizestrant's mechanism of action stops the transcription of ER target genes in wild-type (blue) and mutant (green) ERα, impairing tumor cell proliferation. These properties position camizestrant as a central endocrine therapy partner along with CDK4/6 inhibitors (palbociclib and abemaciclib) in ER⁺ breast cancer. Other signaling pathways are essential to ER⁺ breast cancer proliferation and survival, and contribute to mechanisms of endocrine therapy resistance, including CDK4/6 and PI3K/AKT/mTOR pathways. Inhibitors of these signaling axes are currently approved targeted therapies (everolimus and alpelisib) or under investigation (e.g., capivasertib). *, P < 0.05; **, P < 0.005; ***, P < 0.0005. CAMI, camizestrant; CAPI, capivasertib; CDK, cyclin-dependent kinase; CoA, cytochrome C oxidase assembly; Del, deletion; E2, estradiol; E2F, E2F transcription factor; ERE, estrogen response element; FULV, fulvestrant; m, mutated; MET, metastatic; PALBO, palbociclib; PRIM, primary; RB1, retinoblastoma gene; TGI, tumor growth inhibition.